Treatment of Cancer with Antibodies, and Tumor Targeting of Antibodies
Antibodies have been used to treat cancer for many years. There are currently 5 antibodies approved by the FDA as cancer therapeutics, more than ten in Phase III clinical trials, and several hundred more in Phase II and Phase I trials. Antibodies act to remove cancerous cells through several effector mechanisms. For antibodies that are fully human or chimeric, the Fc portion of the molecule can efficiently activate and interact with the human immune system. By this method, cells can be destroyed by soluble components of the immune system (complement) or through ADCC-mediated cell killing. Additionally, binding of the antibody to the target antigen can initiate a biological response that can lead to apoptosis. Finally, antibody molecules can be used as delivery vehicles to transport therapeutic moieties such as radioisotopes, toxins, or enzymes.
Tumor targeting by antibodies is a complex process that involves circulation and clearance from the bloodstream, diffusion or convection into bulk tumors or micrometastases, binding and release of antigen, and metabolism of antigen/antibody complexes. In order to better understand the processes that influence treatment success, several mathematical models have been developed to analyze the distribution, retention, and removal of full antibody molecules and antibody fragments from solid tumors (Thomas et al., 1989, Sung et al., 1992, van Osdol et al, 1991, Baxter et al., 1995). Most have found there to be an affinity ceiling for uniform penetration and retention. Weinstein and colleagues first proposed the idea of a “binding site barrier” in 1987 (Weinstein et al., 1987). This hypothesis states that antibodies with extremely high affinity would bind antigens at the periphery of the tumor first, which would act as a barrier to further penetration within the tumor. This would result in heterogeneous distribution of the antibody and consequently ineffective treatment. Several additional papers have been published which support this hypothesis with mathematical models (Fujimori et al., 1989, Fujimori et al., 1990, van Osdol et al., 1991, Weinstein and van Osdol, 1992). Subsequent experimental studies have confirmed the existence of this phenomenon (Juweid et al., 1992). Tumor microenvironment can also cause unexpected hindrances. While blood vessels may be more permeable to solute transfer in the tumor (Dvorak et al., 1995, Gerlowski and Jain, 1986), the tumor in general may be less vascularized (Jain, 1999). Tumors also have a high interstitial pressure, which prevents migration of macromolecules into them. Jain et al. have carefully examined the unusual pressure and convection patterns in bulk tumors both experimentally and by mathematical modeling, and have found surprising phenomena such as temporary reverse flows out of tumor tissue.
The first generation of therapeutic antibodies consisted of entire IgG molecules, the format naturally utilized by the mammalian immune system. However, protein engineering technologies have been developed for a wide variety of antibody fragments varying in size and valency. These smaller fragments are more highly diffusible, and therefore penetrate tumor tissue more rapidly. With the advent of directed evolution and display technologies, it is now possible to engineer extremely high affinity antibody fragments. In some instances, experimental biodistribution data indicates limiting returns in targeting as affinity is progressively improved.
Carcinoembryonic Antigen (CEA) and Antibodies thereto in Cancer Treatment
Carcinoembryonic antigen (CEA) has long been identified as a tumor associated antigen (Gold and Freedman, 1965). Originally classified as a protein expressed only in fetal tissue, CEA has now been identified as present in several normal adult tissues. These tissues are primarily epithelial in origin, including cells of the colon, stomach, tongue, esophagus, cervix, sweat glands, and prostrate (Nap et al., 1988, Nap et al., 1992, Prall et al., 1996). Tumors of epithelial origin, as well as their metastases, contain CEA as a tumor associated antigen. While presence of CEA itself does not indicate transformation to a cancerous cell, the distribution of CEA is indicative. In normal tissue, CEA is restricted to the apical surface of the cell (Hammarstrom, 1999). As an example of normal tissue expression, a healthy adult excretes 50–70 mg of CEA in feces per day (Matsuoka et al., 1991). In contrast to normal tissue, cancerous cells tend to express CEA over the entire surface (Hammarstrom, 1999).
Carcinoembryonic antigen is a 180,000 kDa protein with approximately 50% carbohydrate content. It has seven domains, with a single N-terminal Ig variable domain and six domains homologous to the Ig constant domain (Williams and Barclay, 1988). Several distinct epitopes have been identified in CEA (Hammarstrom et al., 1989). Multiple monoclonal antibodies have been raised against CEA for research purposes and as diagnostic tools (Nap et al., 1992). More recently, single chain antibody fragments have been isolated from phage display libraries to be used in radioimmunodetection and radioimmunotherapy (Chester et al., 1994; Osbourn et al., 1999). Of particular interest is the MFE-23 scFv (Chester et al., 1994, WO95/15431, U.S. Pat. No. 5,876,691 to Chester et al.). This antibody fragment has been shown to effectively target colon cancer for radioimmunodetection in viva (Begent et al., 1996). MFE-23 has also been used in radioimmunoguided surgery of colorectal cancer (Mayer et al., 2000). MFE-23 has been produced as a fusion protein to carboxypeptidase G2 (CPG2) and TNFα as well. Both fusions have shown promise in therapy (Chester et al., 2000; Chester et al., 2000, Cooke et al., 2002).